Hydrolysis of steroid semicarbazones



2,961,450 fattest N23132:. 6

2,961,450 7 nYnnoLrsls QF STEROID snMrQa n zoNEs Thomas Eric Holt Bushey Heath, and John Attenburrow, West Harrow, England, assignors to Glaxo Lab- La oratories Limited, .G'reenford, Middlesex, England, a

British company x No Drawing.; Fil ed N ov 10,1 9ss,f s .iso.77z,6so v Claimspi-iority application Great Britain Nov. 14, 1957 s Claims. (Cl. 260 -39745) H This invention is concerned with improvements in or relating to the preparation of steroidicompounds.

r In reactions involving steroids containing one or more keto groups it is sometimes necessary or advisable to protect the het'o groups from attack during the modificatiodof a further part of the steroid molecule. I A

convenient method of protection is to'react the steroid with sen1icarbazide to form the corresponding semicar-- bazone compound. Whilst the keto groups are thus prote'cted, one can often carry out the particular; 'rnodifica V desired, such as for exaniplethje reduction of a keto 7 group inactive t o treatment with semi'carba zid'e, and

thereafter hydrolyse the resultant semi'carbazone to re- ""sto'reiae keto group or -groupsf Thus, for example, in the synthesis of prednisolone,

one may use as intermediate 4:5o6-dihydroco'rtisol acetatepwhich may in turn be prepared from 4:5ix -dihydrocortisone 21- a'cetate by conversion to the corresponding 3:20-bis-semicarbazone, reducing the ll-keto group to an llB-hydroxy group and then hydrolysing the semicarbazone "linkages to' form 4:5-dihydrocortisol. In

like manner, one may convert cortisone Zl-acetate to cortisol.

It is an object of the present invention to provide an. improved procedure for the acid hydrolysis of semi- ;ca rbazone derivatives of keto steroids.

It has now been 'found that the acid hydrolysis of semicarbazone derivatives of steroids is especially conveniently effected in a homogeneousinedium consisting of aqueous tetrahydrofuran and that the resulting keto steroid can be simply recovered'by causing the medium to form two phases, namely an aqueous phase and a tetrahydrofuran phase, the keto steroidbeing largely obtained from the organic phase. Further keto steroid can be obtained by re-extracting the residual aqueous phase with tetrahydrofuran. The separation of the homogeneous re action medium into two phases may be simply effected bya'ddin-g a salt thereto." A

I The principal advantage of this procedure is that one i is able to use the tetrahydrofuran not only to form part of theaetual reaction mixture but also as the extraction solvent. This obviates the use Ora separate and 7,: distinct extraction stage perhaps with a difierent solvent,

the extraction being simply conducted :by mere addition of a salt to the reaction mixture; Tetrahydrofuran is also a very good solvent for steroids and its use as here- V in proposed enables itsfavourable properties to be used to good advantage. Furthermore, the technique of pro tect ion of keto groups' fby semicarbazone formation is of ten us ed whenreact i ve"ketoigroups are to be pro 5 tected during reduction of othe r lceto groups by borohydrides; such reductions are" very conveniently con iz y pr roces sone is aerate conduct such reductions,

sentially' of aqueous"'tetrahydrofuraii, the

. -;ture stirredand the tetrahydro furan l The aqueous layer wasjtwicer g ittetrahydroiuran ,wrththeresult that by the vided -a' process forthe acidlhydr'olysis'pf se'niicarbazone' 1 linkages Lin steroids in which "the hyd'r'dlysi s 'is carried out in solution in a homogeneous :r'nediuni'colris'isting es- 7 siilt'ing keto steroid being recovered by' ca is ing' the me uiiiito separate intotwo' phases fseparatin'g otf theidjrgriic phase and recovering "the keto steroid therefrom.

T e hem en us; stl um.i con nientl fp y adding an aqueous solution of the hydrolysing acid to te ra ydr ea r by addi a id, t t e, u q tetrahydrofuran solution of the semicarba zoneobtained from a previousstep such as reduction Sufficient tetrahydrofuran should be present:in ftlie meaipm to ensure complete solution of the steroid -semic'a'rba'z one and reaction product'; Suitable acids for the hydrolysis inude part cul rlya t on "min ralacis s, era-h eidllp phb ic acids: Su phuric aeid. H

v h P s aee t i 9 thei yem pn ar y ,Qtfvameint e' d oiyslw lsemibatha 9f -ketc pregnane and allopregnan'e" "compounds, which compounds inchideiinpa'rticular the 3'a'ZO-bis-semictrbazones of prednisolone, cortisol, 4:5a-dihydrocortisbl and 4:5}? dihyd'rocortisol land Ith' ZFacylates, e.g.. acetates, thereof.

. In, the applica ion ..of .thepresentjnvention to the I hydrolysis of a" 'semicarbaz'one' of a 20- keto pregnane o1 allopregnane compound; also possessing 'a' 17-hydrox3 group, particularly to the above cited members of tilt series, much improved results have been vfound to b obtained by the use" of sulphuric acid in place of thr previously proposed hydrochloric acid as'tliehydrolysing agent. By this means, a considerable'incriea'se' in yiel is obtained direfit is sa a -ed, 'toi redu' ction in dehydra ion of the 17 -hyd'roXy 'g' The hydrolysis isfconvenientlyeffected at room tern :p'erat'ure by allowinglthe reactionm'ediuin'to stand to i a suflicient time to" eifect substantial hydrolysis of th seniicarbaz one linkages'l After hydrolysis 'is substan ti'ally completethe medium" is conveniently caused t4 separate into two phases by the addition of a'water-solu ble salt. We have found that ammonium sulphate i very effective for this purpose, but other salts, e.g. am monium chloride may also ba -used." The salt shoul' preferablybe' added in such an amount as to effect sub stantially complete separation which may'readily be de termined by. preliminary experiment. Where ammonium sulphate is used la con venient quantity will in genera be from -50%by weight of the amount of water pres eat:

The organic phase recovered is thenwashed (a sat being added to the washing 'solutionstoj'prevent substan tial vup-take of water) and the steroid recovered by suit able means such as'distillationl' The yield of steroid may be,improvedibyextracting th residual aqueous phase with tetrahydrofuran and recoi ering further steroid therefrom after, suitable washing,

In order that the invention'may 'bewellfunderstoot the following examples are given byway' ofillustratio only:

. Exampl 1 n phuric aci 1 I tion inanextremelyconvenient manner.

hydrolysis of the resultant semlcarbazone and extrac- 7 (100 m1) containing ammonium sulphate g.) the with 8% sodium"bic'arbonate solution m1.) 001 taining ammonium sulphate .(25:g':).; 'l heiw'ashed tetra Y Accordingtoathe invention; therefore, thre is p'ro f hydrofuran extracts were dried over anhydrous sodium sulphate and evaporated to dryness yielding crude 4:511.- dihydrocortisol (8.78 g.).

The crude 4:5a-dihydrocortisol (8.78 g.) was treated with acetic anhydride (20 ml.) and pyridine (20 ml.) and heated on a steam bath for 0.5 hour. Excess acetic anhydride and pyridine were removed by distillation under high vacuum and the residue recrystallised from ethyl acetate (about 100 ml.) with charcoal treatment.

Yield of 4:5a-dihydrocortisol acetate:

1st crop: 7.18 g., M.P. 222-225 C., [a] =-|75.2

(1% CHCl 2nd crop: 0.89 g., M.P. 219223 C., [a] =-l-73.8

(1% CHCl The combined crops are equivalent to an overall theoretical yield of 80.7%.

Example 2 4:5a-dihydrocortisol-3:20-bis-semicarbazone (11.5 g.) was hydrolysed and acetylated as in Example 1 with the exception that an hydrolysis time of 3 hours was operative. The 4:5a-dihydrocortisol acetate yield was:

1st crop: 6.84 g., M.P. 222-224 C., [a] =+74.8

(1% CHCl 2nd crop: 0.75 g., M. P. 215-220 C., [a] =-+71.8

(1% CHC13) The combined crops are equivalent to an overall yield of 75.9%.

Example 3 Hydrocortisone-3:ZO-bis-semicarbazone (12.5 g.) dissolved in tetrahydrofuran (290 ml.) and water (90 ml.) was stirred at room temperature with a mixture of sulphuric acid (9 N, 215 ml.) and water (175 ml.). Ammonium sulphate (150 g.) was added and the layers were separated. The tetrahydrofuran layer was stirred for 1 hour with sulphuric acid (2 N, 100 ml.) containing ammonium sulphate (50 g.). and was then washed with saturated sodium bicarbonate solution (100 ml.) containing ammonium sulphate (50 g.). The original layer was stirred for 1 hour with tetrahydrofuran (100 ml.), which was used to re-extract the acid and bicarbonate washes.

The combined tetrahydrofuran solution was dried over sodium sulphate, and the product was isolated by evaporation to dryness and treatment of the residue with ether to give hydrocortisone alcohol Example 4 Cortisone acetate-3:20-bis-semicarbazone (13 g.) was dissolved in tetrahydrofuran (400ml.) and water (200 ml.). Sulphuric acid (9 N, 215 ml.) was added and the solution was stirred at room temperature for 25 minutes. A crystalline solid separated during this period and was collected by filtration.

Paper chromatography indicated that this material was cortisone acetate containing a trace of the 21-alcohol.

The filtrate was stirred with ammonium sulphate (100 g.) and the tetrahydrofuran layer was separated. The solvent layer was washed with sulphuric acid (2 N, 100 ml.) containing ammonium sulphate (50 g.), then with saturated sodium bicarbonate solution (100 ml.) containing ammonium sulphate (50 g.), the aqueous layers being re-extracted with tetrahydrofuran (75 ml.).

The combined tetrahydrofuran solution was dried over sodium sulphate and the product was isolated by evaporation and treatment of the residue with ether to give cortisone acetate (5.1 g.)

iim.= i=+ Example The procedures of Example 1 were repeated using in place of sulphuric acid, 6 N-phosphoric acid. The final total yield of 4:5a-dihydrocortisol acetate was 78.7%.

Example 6 The procedures of Example 1 were repeated using in place of sulphuric acid, 2 N-hydrochloric acid. The hydrolysis time was 1 hour. The total yield of 4:511- dihydrocortisol acetate was 73.5%.

Example 7 Prednisolone 3:20 bis-semicarbazone (1.0 g.) was added to a mixture of tetrahydrofuran (30 ml.) and 5 N sulphuric acid (30 ml.) and stirred for 0.5 hour at room temperature. Ammonium sulphate (8 g.) was added and the mixture stirred and the tetrahydrofuran layer separated. The aqueous layer was re-extracted with tetrahydrofuran (10 ml.). The tetrahydrofuran extracts were washed in turn first with 2 N sulphuric acid (10 ml.) containing ammonium sulphate (5 g.) then with 8% sodium bicarbonate solution (10 ml.) containing ammonium sulphate (5 g.). The washed tetrahydrofuran extracts were evaporated under reduced pressure to about 3 ml. and water (10 ml.) added. The precipitated solid was filtered oil, washed with water (3 ml.) and dried, yielding crude prednisolone (0.08 g.).

We claim:

1. A process for the acid hydrolysis of a steroid blssemicarbazone selected from the group consisting of a bis-semicarbazone of a 3:20-diketosteroid of the pregnane series and a bis-semicarbazone of a 3:20-diketosteroid of the allopregnane series, comprising contacting said semicarbazone with a strong mineral acid in a homogeneous liquid medium consisting of aqueous tetrahydrofuran to hydrolyze the semicarbazone linkages; adding a watersoluble salt to said medium to effect separation thereof into an organic phase and an aqueous phase; separating said organic phase from said aqueous phase; and recovering the hydrolyzed steroid from said organic phase.

2. The process of claim 1 in which said hydrolyzed steroid is recovered from the organic phase by distilling oil the tetrahydrofuran.

3. The process of claim 1 in which said bis-semicarbazone is the 3:20-bis-semicarbazone of prednisolone.

4. The process of claim 1 in which said bis-semicarbazone is the 3:20-bis-semicarbazone of prednisolone-Zlacetate.

5. The process of claim 1 in which said bis-semicarbazone is the 3 :20-bis-semicarbazone of cortisol.

6. The process of claim 1 in which said bis-semicarbazone is the 3:20-bis-semicarbazone of cortisol Zl-acetate.

7. The process of claim 1 in which said bis-semicarbazone is the 3:20-bis-semicarbazone of 4:5a-dihydrocortisol.

8. The process of claim 1 in which said bis-semicarbazone is the 3:20-bis-semicarbazone of 4:5a-dihydrocortisol ZI-acetate.

9. The process of claim 1 in which said bis-semicarbazone is the 3:20-bis-semicarbazone of 4:5}9-dihydrocortisol.

10. The process of claim 1 in which said bis-semicarbazone is the 3:20-bis-semicarbazone of 4:5;9-dihydrocortisol ZI-acetate.

11. The process of claim 1 in which said strong mineral acid is sulfuric acid.

12. The process of claim 11 in which said water-soluble salt is ammonium sulfate.

13. The process of claim 12 in which the ammonium sulfate added is from 25-50% by weight of the water present in said medium.

Kapp et a1. Nov. 14. 1950 Day Feb. 12, 1957 

1. A PROCESS FOR THE ACID HYDROLYSIS OF A STEROID BISSEMICARBAZONE SELECTED FROM THE GROUP CONSISTING OF A BIS-SEMICARBAZONE OF A 3:20-DIKETOSTEROID OF THE PREGNANE SERIES AND A BIS-SEMICARBAZONE OF A 3:20-DIKETOSTEROID OF THE ALLOPREGNANCE SERIES, COMPRISING CONTACTING SAID SEMICARBAZONE WITH A STRONG MINERAL ACID IN A HOMOGENEOUS LIQUID MEDIUM CONSISTING OF AQUEOUS TETRAHYDROGURAN TO HYDROLYZE THE SEMICARBAZONE LINKAGES, ADDING A WATERSOLUBLE SALT TO SAID MEDIUM TO EFFECT SEPARATION THEREOF INTO AN ORGANIC PHASE AND AN AQUEOUS PHASE, SEPARATING SAID ORGANIC PHASE FROM SAID AQUEOUS PHASE, AND RECOVERING THE HYDROLYZED STEROID FROM SAID ORGANIC PHASE. 